Materials and Methods

Animals—Fifty-eight pregnant (approx 2 to 3 weeks of gestation) Hartley guinea pigs were obtained from a commercial source^a for use in the study. Seventeen guinea pigs were used in a preliminary IP challenge experiment. Mean weight at time of arrival for these guinea pigs was 865 g. Subsequently, 41 pregnant guinea pigs were used in an oral challenge experiment. Mean weight at time of arrival for these guinea pigs was 797 g. At arrival, a rectal swab specimen was obtained from each guinea pig, placed in transfer medium, and plated for Campylobacter culture.

Guinea pigs were housed in standard metal cages with wood chip bedding and fed a pelleted commercial diet formulated for guinea pigs. Animals were housed in groups (4 or 5 guinea pigs/cage for the IP challenge exposure and 3 or 4 guinea pigs/cage for the oral challenge exposure) and allowed to acclimate to their environment for 7 days prior to inoculation. During this time, weights were recorded daily and guinea pigs were marked with black hair dye to provide unique identification for each animal. For the oral challenge exposure, treatment groups were housed in separate banks of cages; strict hygienic procedures were used between handling of groups to prevent cross-contamination. All procedures were approved by the Institutional Animal Care and Use Committee at Iowa State University.

Campylobacter strains—Campylobacter jejuni isolates were used in challenge exposures. The IA 3902 isolate was cultured from an aborted ovine fetus. This tetracycline-resistant strain of C jejuni is the predominant genetic clone associated with abortion in sheep, as determined by pulsed-field gel electrophoresis, multilocus sequence typing, and cmp gene sequence typing.⁸ The OF48 isolate was cultured from feces of a sheep that did not have clinical signs (abortion or diarrhea). The NCTC 11168 isolate^b is from humans and has been completely sequenced.¹⁸ Fresh bacterial cultures were obtained following 24 hours of growth on MH agar in anaerobic jars under microaerobic conditions (5% oxygen, 10% carbon dioxide, and 85% nitrogen) at 42°C. These cultures were collected and placed in MH broth, diluted to desired concentrations on the basis of optical density, and then used as inocula in the challenge experiments. The final number of organisms in each suspension was determined by counting the number of viable CFUs.

IP challenge exposure—Two groups of guinea pigs were used in a preliminary experiment. The first group (which comprised 5 guinea pigs) served as sham-inoculated control animals and were inoculated IP with 1.0 mL of MH broth IP, whereas the second group (which comprised 12 guinea pigs) was inoculated IP with 1.0 mL of 1.5 × 10⁵ CFUs of C jejuni IA 3902/mL of MH broth. This concentration was chosen on the basis of reports^12,13 in which pregnant guinea pigs were challenge exposed with C jejuni via IP inoculation. Guinea pigs in both groups were inoculated by use of 1-mL tuberculin syringes with a 26-gauge, 3/8-inch needle.

Oral inoculation—Four groups of guinea pigs were used in the oral challenge exposure experiment. The first group (n = 5 guinea pigs) served as sham-inoculated control animals and received 1.0 mL of MH broth. The second group (n = 12 guinea pigs) received 1.0 mL of 5.5 × 10⁸ CFUs of C jejuni OF48/mL of MH broth. The third group (n = 12 guinea pigs) received 1.0 mL of 5.9 × 10⁸ CFUs of C jejuni NCTC 11168/mL of MH broth. The fourth group (n = 12 guinea pigs) received 1.0 mL of 6.0 × 10⁸ CFUs of C jejuni IA 3902/mL of MH broth. These doses were chosen to approximate those used in other experiments in which C jejuni was administered orally to pregnant ferrets¹⁹ and guinea pigs.¹⁷ Guinea pigs in all 4 groups were inoculated orally via a curved, 18-gauge, 3-inch, stainless-steel feeding needle. Food was withheld from the guinea pigs for 12 hours prior to inoculation to diminish the amount of food retained in the oral cavity.

Monitoring, euthanasia, necropsy, and collection of samples—After inoculation, guinea pigs were weighed daily by use of a digital gram scale and were observed at least twice daily for signs of abortion or impending abortion. These signs included vaginal bleeding, expelled fetuses or fetal membranes, and sudden substantial weight loss (> 10% in 24 hours). Once a guinea pig had aborted or an impending abortion was identified, that animal was euthanized via IP injection of sodium pentobarbital (approx 150 mg/kg) and immediately necropsied. All guinea pigs that had not aborted were euthanized 21 days after inoculation and necropsied.

Necropsy of guinea pigs involved inspection for gross lesions and collection of samples for bacterial culture and histologic examination. Rectal swab specimens for Campylobacter culture were obtained by use of sterile minitip cotton swabs^c immediately after euthanasia. Other samples harvested for Campylobacter culture included heart blood, bile, and uterine tissue from each dam and placenta, liver, and lung tissues from each fetus. Pooled fetal stomach contents were collected only during the oral challenge portion of the study. Heart blood, bile, and fetal stomach contents were collected by use of a sterile tuberculin syringe with a 26-gauge, 3/8-inch needle. Samples of uterine, placental, and pooled fetal liver and lung tissues were placed in separate sterile plastic bags^d (IP challenge) or separate sterile Petri plates (oral challenge). All samples were refrigerated until immediately prior to culture, with bacterial culture performed on most samples on the day of collection.

Samples collected for histologic examination included liver, gallbladder, and uterine tissues for each dam and placenta, liver, and lung tissues for each fetus. Samples obtained for histologic examination were placed in neutral-buffered 10% formalin for 24 hours, embedded in paraffin, and processed routinely for H&E staining.

Campylobacter culture and semiquantitative enumeration of C jejuni from necropsy samples—A few drops of each fluid sample (blood, bile, or fetal stomach contents) were placed directly onto culture media and streaked by use of sterile cotton swabs. Rectal swab specimens were streaked directly onto culture media. Uterine tissues, pooled fetal liver and lung tissues, and placental tissues were minced with a scalpel or scissors; then, swab specimens were obtained from the minced tissues by use of a sterile cotton swab and streaked onto culture media. All samples were spread onto MH agar containing a Campylobacter selective supplement^e (polymyxin B, rifampicin, trimethoprim, and cycloheximide) and a Campylobacter growth supplement^f (sodium pyruvate, sodium metabisulfite, and ferrous sulfate) and were incubated for 48 hours in anaerobic jars under microaerobic conditions at 42°C. After incubation, Campylobacter-like colonies on each plate were counted to determine the number of CFUs in each sample.

Histologic scoring of uterine lesions—Endometrial inflammation was evaluated by calculating the mean number of leukocytes in 10 hpfs (40X magnification). Uterine lesions were scored in each hpf by use of the following criteria: 0 = inflammatory infiltrate was not identified, 1 = < 10 leukocytes/hpf, 2 = 10 to 20 leukocytes/hpf, and 3 = > 20 leukocytes/hpf. An additional 0.5 points was added to the score when the inflammation extended into the underlying myometrium.

Immunohistochemical analysis—Placental samples were collected at necropsy and embedded in paraffin, as described previously. Tissues were sectioned at a thickness of 3 μm, mounted on aminoalkylsilanecoated glass slides, and placed in an oven at 56°C for 2 hours. Sections were routinely deparaffinized in xylene and rehydrated in graded alcohol solutions and water baths. Endogenous peroxidase inhibition was achieved by immersion (2 immersions; 10 min/immersion) in baths of 3% H₂O₂ in water. Sections were incubated with 0.1% protease in a Tris buffer (pH, 7.6) at 37°C for 15 minutes. Slides were rinsed 3 times in PBS solution and then were placed in an automated cell staining system.^g To inhibit nonspecific binding, sections were incubated in 10% neutral goat serum at 22°C for 20 minutes. The primary antibody, which was directed against the major outer membrane protein of C jejuni, was prepared as described in another study.²⁰ This antibody was used at a dilution of 1:300; slides were incubated at 22°C for 60 minutes followed by rinsing in a bath of PBS solution for 5 minutes. A commercially available biotinylated secondary antibody^h was used at a dilution of 1:80; slides were incubated at 22°C for 15 minutes followed by rinsing in a bath of PBS solution for 5 minutes. Sections were then incubated with horse radish peroxidase–streptavidin conjugateⁱ at 22°C for 15 minutes followed by rinsing in a bath of PBS solution for 5 minutes. The final reaction was developed by use of a commercial chromogen.^j Sections were rinsed and routinely counterstained with Shandon Harris hematoxylin^k and Scott's tap water. Sections were dehydrated through graded alcohol and xylene solutions prior to mounting. Positive control samples were obtained from paraffin blocks of ovine placental tissues that had positive results when cultured for C jejuni IA 3902. Negative control samples consisted of sections from culture-negative tissues obtained from nonaborting guinea pigs inoculated with C jejuni IA 3902 and from sham-inoculated control guinea pigs.

Statistical analysis—A commercial statistical software package^l was used to perform 1-way ANOVAs to detect differences among groups. Abortion rates, mean uterine scores, necrosuppurative placentitis, and random necrotizing hepatitis were compared. Results were considered significant at values of P ≤ 0.05.

Results

IP challenge exposure—All rectal swab specimens collected before challenge exposure had negative results when cultured for Campylobacter spp. Ten of 12 guinea pigs inoculated with IA 3902 aborted, with 8 guinea pigs aborting 2 or 3 days after inoculation and the remaining 2 guinea pigs aborting 8 days after inoculation. Necropsy revealed that the 2 guinea pigs that did not abort were not pregnant; 1 of these guinea pigs had a teratoma of the left ovary. For the control group, none of the 5 guinea pigs aborted and necropsy revealed that all 5 were pregnant.

Calculation of the abortion rate excluded the 2 guinea pigs that were not pregnant. Thus, the mean abortion rate for guinea pigs inoculated IP with IA 3902 (100%) was significantly (P < 0.001) higher than the mean abortion rate for the control guinea pigs (0%).

Oral challenge exposure—All rectal swab specimens collected before challenge exposure had negative results when cultured for Campylobacter spp. Six of 12 guinea pigs inoculated with IA 3902 aborted; 4 guinea pigs aborted between 4 and 6 days after inoculation, and the other 2 guinea pigs aborted between 7 and 10 days after inoculation. None of the guinea pigs in the remaining groups aborted. Necropsy revealed that 2 guinea pigs in each of the IA 3902, NCTC 11168, and OF48 groups and 1 guinea pig in the control group were not pregnant; we determined that these guinea pigs had not been pregnant during the study on the basis of a lack of appreciable weight gain. These guinea pigs were excluded from the calculations for abortion rate.

Mean abortion rates differed significantly (P < 0.001) among groups (Table 1). The mean abortion rate for guinea pigs inoculated with IA 3902 (6/10) was significantly (P < 0.001) higher than the rate for any of the other 3 groups (0/10, 0/10, and 0/4 for guinea pigs inoculated with NCTC 11168, OF48, or the sham inoculation, respectively). In addition, necropsy revealed that 1 of the 4 pregnant, nonaborting guinea pigs from the IA 3902 group had histologic and bacteriologic evidence of fetal infection.

Gross liver lesions consisted of multiple, variably sized, round, white foci; they were identified in 9 guinea pigs inoculated with IA 3902 and 3 guinea pigs inoculated with NCTC 11168. The predominant histopathologic liver lesion was random, multifocal to coalescing hepatitis, which was detected in decreasing frequency in the IA 3902 (11/12), NCTC 11168 (8/12), and OF48 (4/12) groups; this lesion was not detected in the control group (0/4). Inflammatory foci were variably sized, and the associated infiltrate differed depending on the interval between challenge exposure and necropsy. In guinea pigs that aborted within a few days after challenge exposure, inflammatory foci were primarily neutrophilic. As the interval between challenge exposure and abortion increased, the infiltrate became dominated by lymphocytes, plasma cells, and macrophages.

Mean incidence of multifocal random hepatitis differed significantly (P < 0.001) among groups. The mean incidence of random hepatitis was significantly (P < 0.001) higher in the IA 3902 group, compared with the mean incidence for the other 3 groups. The mean incidence of random hepatitis was significantly (P = 0.004) higher in the NCTC 11168 group, compared with the mean incidence for the control group, but did not differ significantly (P = 0.092) from the mean incidence for the OF48 group. Mean incidence or random hepatitis for the OF48 group did not differ significantly (P = 0.136) from the mean incidence for the control group. Mild periportal infiltrates of lymphocytes and plasma cells were detected in most of the samples from all groups (Table 1). Additionally, congestion or edema of the gallbladder was evident in 5 guinea pigs of the IA 3902 group, 5 guinea pigs of the NCTC 11168 group, 3 guinea pigs of the OF48 group, and 1 guinea pig of the control group.

Uterine lesions were variable and consisted of suppurative endometritis, metritis, and hemorrhage of varying severity. Uterine edema was evident in guinea pigs from all the IA 3902, NCTC 11168, and OF48 groups but was most severe in those from the IA 3902 group. In general, uterine inflammatory scores were highest in guinea pigs inoculated with IA 3902 (mean, 2.3; range, 1.0 to 3.5), with lower scores in a few guinea pigs of the OF48 (mean, 0.4; range, 1.0 to 2.5) and control (mean, 0.5; range, 1.0 to 1.5) groups and no metritis detected in the NCTC 11168 group.

Mean uterine scores differed significantly (P < 0.001) among groups (Table 1). Mean uterine score for the IA 3902 group was significantly (P < 0.001) higher than the mean uterine score of any of the other 3 groups, whereas the mean uterine scores for the NCTC 11168 and OF48 groups did not differ significantly (P = 0.294 and P = 0.792, respectively) from the mean uterine score for the control group.

Microscopic placental lesions consisted of a combination of hemorrhage, suppurative inflammation, and necrosis (Figure 1). Placental lesions were evident in 8 guinea pigs inoculated with IA 3902 (including all 6 that aborted) and in 1 guinea pig inoculated with NCTC 11168. These changes were detected in both the maternal and fetal portions of the placenta, with severity of the lesions increasing with an increase in the number of days after inoculation.

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Photomicrographs of sections of placental tissues obtained from guinea pigs inoculated with Campylobacter jejuni IA 3902. A—Notice the necrosuppurative placentitis and hemorrhage at the junctional zone (JZ) between the subplacenta (SP) and main placenta (MP) and marked vasculitis of the adjacent uteroplacental artery (UA). H&E stain; bar = 200 μm. B—Photomicrograph of the same section of placental tissue in panel A after immunohistochemical staining for C jejuni. Notice the C jejuni organisms (red stain) located within inflammatory cells and in extracellular spaces in the junctional zone. Primary antibody was directed against the major outer membrane protein of C jejuni, followed by a biotinylated secondary antibody and incubation with horse radish peroxidase–streptavidin conjugate and developed by use of a commercial chromogen. Counterstained with Shandon Harris hematoxylin and Scott's tap water; bar = 200 μm. C—Photomicrograph of a section of guinea pig placental tissue after immunohistochemical staining for C jejuni. Notice the C jejuni organisms (red stain) within the trophoblasts and syncytial streamers (SS) of the subplacenta. Bar = 200 μm. D—Photomicrograph of a uteroplacental artery (UA) from a guinea pig that aborted 8 days after inoculation. Notice the marked vasculitis and abundant perivascular C jejuni organisms (red stain). Immunohistochemical stain for C jejuni; bar = 200 μm.

Citation: American Journal of Veterinary Research 70, 10; 10.2460/ajvr.70.10.1269

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Mean incidence of necrosuppurative placentitis differed significantly (P < 0.001) among groups (Table 1). Mean incidence of placentitis in the IA 3902 group (8/10) was significantly (P < 0.001) higher than the mean incidence for the NCTC 11168 (1/10), OF48 (0/10), and control (0/4) groups; mean incidence for the NCTC 11168 and OF48 groups did not differ significantly (P = 0.569 and P = 1.000, respectively), compared with the mean incidence for the control group.

High numbers of C jejuni were cultured from 7 of 10 placental samples, 6 of 10 fetal liver or lung samples, and 6 of 10 uterine samples in the IA 3902 group. Moderate numbers of C jejuni were cultured from 10 of 12 rectal swab specimens in the IA 3902 group, and low numbers of C jejuni were generally cultured from 5 of 12 heart blood samples, 4 of 9 fetal stomach contents samples, and 2 of 12 bile samples in the IA 3902 group. Low to moderate numbers of C jejuni were cultured from 9 of 12 rectal swab specimens from the NCTC 11168 group and 6 of 12 rectal swab specimens from the OF48 group. Campylobacter jejuni was not cultured from any other tissues in these groups or from any samples collected from the control group. The percentage of samples with positive results for culture of C jejuni were summarized (Figure 2). The source and number of CFUs for bacterial cultures of guinea pigs inoculated with IA 3902 were also summarized (Table 2).

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Percentage of samples with positive results when cultured for C jejuni. Samples were obtained during necropsy of guinea pigs after oral inoculation with C jejuni strain IA 3902, NCTC 11168, or OF48 or a sham inoculum. Necropsies were performed at the time of abortion or at 21 days after inoculation in guinea pigs that did not abort.

Citation: American Journal of Veterinary Research 70, 10; 10.2460/ajvr.70.10.1269

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Immunohistochemical analysis—Immunohistochemical analysis for C jejuni was performed on all available placental tissues from guinea pigs that aborted during the study. Staining for C jejuni was not detected in any negative control samples or culture-negative aborted tissues. In guinea pigs inoculated IP with IA 3902, staining for C jejuni was evident in 5 of 6 culturepositive samples (placental tissues or fetal tissues when placental samples were not available for culture). In guinea pigs inoculated orally with IA 3902, staining for C jejuni was evident in 5 of 7 culture-positive samples. Campylobacter jejuni organisms were identified within the cytoplasm of subplacental trophoblasts, within phagocytes, and in extracellular spaces surrounding trophoblasts in areas of placentitis and necrosis (Figure 1). Organisms were most prevalent at the periphery of the subplacenta within the junctional zone. Multifocal intracellular organisms were also detected within the deeper portions of the endometrium and myometrium and surrounding the uteroplacental arteries; however, this distribution was limited to samples from guinea pigs that aborted at longer intervals after inoculation.

Discussion

Campylobacteriosis has been described as the most common cause of infectious abortion in sheep in the United States,² and C jejuni has become the predominant species.^7,8 Pulsed-field gel electrophoresis, multilocus sequence typing, and cmp gene sequence typing of C jejuni isolates from multiple farms in multiple states during multiple lambing seasons have revealed that most of these isolates are of a single clone, which is represented by C jejuni IA 3902.⁸ In the guinea pigs of the study reported here, we identified several unique features of C jejuni IA 3902, when compared with those of a sheep commensal fecal isolate (C jejuni OF48) and a fully sequenced human isolate (C jejuni NCTC 11168).

Campylobacter jejuni IA 3902 was uniquely proficient in its ability to induce abortion. In the preliminary experiment with IP challenge exposure, which was intended to verify the abortifacient potential of this isolate, all of the pregnant guinea pigs aborted. After oral inoculation, C jejuni IA 3902 was the only isolate that induced abortion (6/10 pregnant guinea pigs aborted). Had the study been extended a few additional days, the abortion rate after oral inoculation would likely have been even higher for IA 3902 because placentitis and fetal infection were identified in 1 guinea pig that had not yet aborted when the study was terminated. This is in contrast to results of a study¹⁷ in which 4 pathogenic human isolates of C jejuni were evaluated in pregnant guinea pigs (abortion rates for those isolates ranged from 0% to 67% following IP inoculation and 0% following oral inoculation). To our knowledge, the study reported here is the first to reveal that oral inoculation of C jejuni can be effective in inducing abortion in pregnant guinea pigs. These results indicated that C jejuni IA 3902 was highly abortifacient and that pregnant guinea pigs were an effective method for evaluating the pathogenicity of this isolate for a natural route of infection (oral) or after IP inoculation.

The pathogenesis of Campylobacter spp that induce abortion when acquired by a nonvenereal route of transmission entails oral exposure, intestinal colonization, bacterial invasion, bacteremia, and infection of the fetoplacental unit.^1,21–23 Sheep appear to be at increased risk for developing campylobacteriosis because healthy sheep often harbor Campylobacter spp within the intestines and gallbladder.^1,24 In a recent study,²⁵ Campylobacter spp were identified in 49.5% of intestinal, gallbladder, and fecal samples obtained from healthy sheep. The pathogenic mechanisms used by C jejuni in the evasion of the physical and immunologic barriers in the intestines have been extensively studied and have been described elsewhere.^26,27 In the study reported here, semiquantitative cultures of fecal samples revealed that all 3 strains of Campylobacter organisms were capable of colonizing the intestinal tract after oral administration, with C jejuni IA 3902 cultured in the greatest number of samples (10/12 [83%]) collected during necropsy, followed by C jejuni NCTC 11168 (9/12 [75%]) and OF48 (6/12 [50%]).

Once the intestines have been colonized, abortifacient Campylobacter spp must breach the intestinal epithelium and induce bacteremia. Bacteremia attributable to C jejuni has been described,^28–30 and results of 1 study³¹ indicated that certain strains of C jejuni have an enhanced ability to induce bacteremia. Because of the stress associated with blood collection and the limited blood volume that can be harvested from guinea pigs on a daily basis, the magnitude and duration of bacteremia were not directly assessed in the present study, and an indirect gauge of systemic infection was used to estimate the incidence of bacteremia. Acute hepatitis has been reported as a sequela to bacteremia attributable to C jejuni in humans,³² and liver biopsy specimens from a patient with acute hepatitis and concurrent enteritis attributable to C jejuni revealed nonspecific reactive hepatitis with focal necrosis.³³ Additionally, multifocal random hepatitis is a consistent necropsy finding secondary to bacteremia or septicemia for numerous organisms, including host-adapted Salmonella spp³⁴ such as Salmonella enterica serovar Dublin in cattle and Salmonella enterica serovar choleraesuis in swine. In the study reported here, the identification of multifocal, random hepatitis was used as indirect evidence of systemic infection. This lesion was identified in 23 of 36 orally inoculated guinea pigs, including 11 of 12 guinea pigs inoculated with C jejuni IA 3902 and 8 of 12 guinea pigs inoculated with C jejuni NCTC 11168. The significantly higher rate of hepatitis in both groups indicated that each strain likely induced a degree of bacteremia, and it therefore seems likely that the differences in abortion rate were attributable to a mechanism other than the ability of these organisms to breach the intestinal epithelium and induce bacteremia.

Predilection for the fetoplacental environment is a common attribute of abortifacient agents, and it appears that the greatest difference between C jejuni IA 3902 and the other evaluated isolates was its affinity for the fetoplacental unit. All 3 isolates were able to colonize the intestines, and there was evidence that both C jejuni IA 3902 and C jejuni NCTC 11168 induced a substantial bacteremia after oral inoculation, yet only IA 3902 appeared to be able to colonize the placenta, infect trophoblasts, and induce abortion. Guinea pigs inoculated with C jejuni IA 3902 consistently developed endometritis and placentitis with high numbers of C jejuni (> 1,000 CFUs) recovered from uterine, placental, and pooled fetal tissues, all of which provided evidence for an apparent tropism of IA 3902 for the fetoplacental unit.

Evaluation of our results indicated that immunohistochemical analysis was less sensitive than microbial culture as a means of detecting C jejuni in tissues, which was consistent with results in other studies.^35,36 However, immunohistochemical analysis was extremely useful in characterizing the distribution of organisms in placental tissues. Campylobacter jejuni IA 3902 appeared to localize at the periphery of the subplacenta near its junction with the main placenta early during the course of infection and was evident within inflammatory cells and syncytiotrophoblasts of the subplacenta. This localization appears logical because this is the site of placental vascular invasion³⁷ and would be the first location reached during episodes of bacteremia. Additionally, guinea pigs infected for longer periods before they aborted often had immunohistochemical evidence of abundant pericellular and intracytoplasmic organisms associated with the periarterial trophoblasts in the decidua. These data suggested that trophoblasts of subplacental origin were preferentially infected, whereas those of the main placenta were spared. It is worth mentioning that there are considerable differences between the placentation of guinea pigs and ewes. Guinea pigs have a discoid, labyrinthine, hemochorial placenta with extensive trophoblast invasion of the decidua and uterine arterial walls.^38,39 In contrast, sheep have a cotyledonary, epitheliochorial placenta and lack trophoblast invasion of uterine blood vessels.³⁹ Although there are noticeable differences in placentation between these 2 species, our immunohistochemical findings of organisms within trophoblasts, leukocytes, and perivascular tissues were consistent with those in another study⁴⁰ of placentas from aborting sheep infected with C jejuni. Together, these findings suggest that pathogenic C jejuni strains are capable of infecting trophoblasts regardless of placentation type, and this provides further supporting evidence for the appropriateness of the use of pregnant guinea pigs as a method for evaluation of Campylobacter-induced abortion. To the authors' knowledge, the information reported here was the first description of the localization and distribution of Campylobacter spp within the placenta of aborting guinea pigs.

In this study, we determined that C jejuni IA 3902 was a unique, highly abortifacient strain with the ability to colonize the intestines, cause systemic infection, and induce abortion as a result of its affinity for the fetoplacental unit. Although we confirmed the highly pathogenic nature of IA 3902 in pregnant guinea pigs, additional in vivo studies in sheep are required to determine the degree of virulence in pregnant ewes. Campylobacter jejuni IA 3902 appears to be a valuable strain for studying the pathogenesis of Campylobacter-induced abortion. To facilitate the study of pathogenic mechanisms, the genome of IA 3902 is being sequenced by our laboratory group, and we will compare it with a fully sequenced, nonabortifacient strain (NCTC 11168) to attempt to identify unique virulence traits of this abortifacient clone. Results also revealed that pregnant guinea pigs were useful for studying the pathogenesis of abortion attributable to this unique Campylobacter strain. Pregnant guinea pigs have been used to assess the efficacy of vaccines against C fetus,^14–16 and given the recent shift from C fetus to predominantly C jejuni causing abortions in sheep,⁸ there is an urgent need to explore the development of a vaccine against C jejuni for the control of this emergent, highly pathogenic strain.